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Chapter 7 : Porin Regulon of Escherichia coli
In Escherichia coli, the two porin proteins OmpF and OmpC form pores in the outer membrane that allow for the passive diffusion of small hydrophilic molecules across this hydrophobic barrier. Studies using gene and operon fusions to both ompF and ompC revealed that regulation of porin expression occurs at the transcriptional level. This work, combined with additional genetic analysis, led to the proposal of an early model to explain porin regulation. Importantly, strains merodiploid for envZ473 and either envZ247 or envZ250 exhibit intermediate levels of porin expression that are comparable with that normally seen in low osmolarity. OmpR can activate transcription of both the ompF and ompC genes and can also repress transcription of ompF. To regulate expression of the porin genes, OmpR must interact with regulatory regions of the ompF and ompC promoters in a manner that results in the activation and/or repression of transcription. Much study has been focused on elucidating the following: the regions of DNA to which OmpR binds and the nucleotide sequence of these regions; how OmpR recognizes these regions; and finally, how these OmpR-promoter interactions affect DNA topology, functional interactions with the polymerase, and ultimately transcriptional activation and/or repression. Importantly these conditions mimic those found within the bodies of animals. In external environment surroundings, nutrients are scarce, and the slightly wider OmpF channel enables their more efficient uptake. The smaller, more protective OmpC porin is no longer required in this very dilute environment.
Domain structure of OmpR and EnvZ. The sensor, EnvZ, is located in the inner membrane with its N-terminal domain in the periplasm and its C-terminal domain in the cytoplasm. It possesses two transmembrane segments extending from 16 to 46 (TM1) and from 163 to 179 (TM2).The N-terminal domain of EnvZ monitors the osmolarity, and this information is transduced across the inner membrane to the C-terminal cytoplasmic domain. The C-terminal portion of EnvZ relays this information to the response regulator, OmpR, via kinase and phosphatase activities. OmpR is a cytoplasmic protein directly responsible for regulating transcription of the ompF and ompC genes. The N-terminal half of OmpR is the phosphorylation domain, containing the acidic pocket and the presumed site of phosphorylation (Asp-55), and the C-teminal portion of OmpR is the DNA binding domain.
Signaling states of EnvZ. EnvZ must assume different signaling states under varying osmotic conditions. This is accomplished by regulating the ratio of kinase to phosphatase activities such that a phosphatase-dominant state is present in low osmolarity whereas a kinase-dominant state predominates in high osmolarity. The ratio of kinase to phosphatase activities could be regulated by altering either or both enzymatic activities. One possibility is that the ratio is regulated by controlling the relative exposure of the autophosphorylation site, thereby regulating the rate of autophosphorylation (see text for further details).
Nature of the internal signal. (Top) OmpR assumes different roles under varying osmotic conditions. In low osmolarity (thick line), OmpR functions as a transcriptional activator of ompF, whereas in high osmolarity (thin line), OmpR functions to repress ompF transcription and activate ompC transcription. (Bottom) The distinction between the OmpR in low versus high osmolarity is thought to be a difference in the concentration of OmpR-phosphate. Low concentrations of OmpR favor transcriptional activation of ompF, whereas higher concentrations of OmpR-phosphate repress ompF transcription and activate ompC transcription.
Porin promoters. OmpR-mediated transcriptional regulation of the porin genes involves extensive regions of promoter DNA. The regions required for positive and negative regulation are indicated above the promoters by (+) and (−), respectively. The regions protected by OmpR in vivo and in vitro are also indicated (see key).